Study On Enzymatic Extraction For Tremella Fuciformis Polysaccharides And Its Biological Activity

Tremella fuciformis,known as white fungus,snow fungus,or chrysanthemum fungus,is an edible fungus of great value for further research and development.Tremella fuciformis polysaccharide(shortening TFP),the principal active constituent,becomes a research hotspot due to its physiological function on improving body immunity against tumor and virus,lowering blood glucose and blood lipid.However,compared with the developed countries like Europe and America,tremella in China is mainly used as food and the research on TFP started relatively late.In the extraction of TFP exists some problems such as complex process,low extraction rate and purity,and expensive costly.And the contradiction between supply and demand occurs due to the demand for high-purity TFP at home and abroad.Fujian Provice is rich of tremella resources.Therefore,the study of the extraction and purification of TFP is of practical significance and application value for the development of a simple,economical and efficient method to extract and purify polysaccharides of tremella fuciformis.This paper focused on TFP extraction,separation and its purification,and physiological activity.The main contents and results of this study were as follows:(1)Extraction of TFP by Enzymolysis combined with High temperature cooking methodThe dry tremella fruit was treated with high pressure cooking after soaking,pruning,draining with pure water,and then the TFP would be extracted after homogenating,enzymolysis,alcohol precipitation and centrifugal freeze-drying.The single factor experiment was conducted to expore the effects of temperature,time,solid-liquid ratio during high temperature treatment,on the extration rate of TFP.The parameters of high temperature treatment was optimized by Box-Behnken experiment follows:temperature 110 ??time 120 min?solid-liquid ratio 1:90.After the process of high temperature and colloid grinding,the effects of single factors in the stage of enzymolysis,namely cellulase dosage,pectinase dosage and the time and temperature of enzyme digestion,on the extraction rate of TFP were studied.Then the orthogonal experimental design was employed to optimize such the above-mentioned process,and the optical technological parameters were obtained as follows:cellulose enzyme dosage 1.0%?pectinase dosage 0.8%?the time 120 min.the temperature 55?,the extraction ratio was 39.28%.(2)The Separation and Purification of TFPThe separation and purification for TFP was the basis to study its physiological activity.Two simple constituents TFP 1-1 and TFP2-2 were isolated and purified by treatments namely deproteinzation,ion-exchange column chromatography DEAE-Cellulose,Sephadex G-200 gel filtration chromatography and dialysis.And then the deproteinzation rate and loss rate of TFP were selected as the comprehensive evaluation index.To compare with the efficiency of TCA,sevage,hydrochloric acid methods,the result showed that the method of enzymolysis with sevage was tested to be the optimal method.Under the the conditions 65 ?,pH 6.5,treating time 120 min and papain addition 1%.After protein removed by sevege method for three times,the deproteinzation TFP reached up to 85.1%,and the loss rate was 14.2%.(3)The Study on the Physiological Activity of TFPBy comparison the effects on · OH clearance between the TFP,deproteinized TFP treatment,the result showed that the radicals scavenging ability of both kinds TFP was significantly(p<0.01)less than that of Vc.While the radicals scavenging ability of TFP was significantly(p<0.01)less than that of deproteinized.In the ABTS+ system,TFP also indicated good clearance effect.The IC50 of the deproteinzed and the untreated TFP was 2.32mg/mL and 3.94mg/mL respectively in ABTS+ clearance.However,in the system of DPPH,the TFP without deproteinzation did not perform so well as the deproteinzed in the removal of DPPH free radicals(p<0.01).Next,the paper had discussed the anti-tumor function of TFP and found out that the TFP in combination with cisplatin had positive effect on suppressing the growth of HepG2 hepatoma carcinoma cells,in which,the suppression rate of untreatedTFP reached 71.8%when combined with cisplatin,while the deproteinzed showed 63.2%.The conclusion was drawn that the TFP without deproteinzation was more effective than the deproteinzed in the supression of cell growth when combined with cisplatin.